Project Summary There is compelling evidence to support the hypothesis that the underlying cause of aging is the cell autonomous, time-dependent accumulation of stochastic damage to cells, organelles and macromolecules. However, it is also clear from parabiosis, plasma transfer, and cell ablation studies that cell non-autonomous mechanisms play important roles in modulating degenerative changes that arise as the consequence of spontaneous, stochastic damage with age. For example, GDF-11 and oxytocin have been implicated as having tissue-specific anti- geronic effects whereas Wnt, 2-microglobulin and CCL11 have been implicated as blood-borne, tissue or cell- type specific pro-geronic factors. In addition, senescent cells, which accumulate in tissues with age, can display a senescence-associated secretory phenotype (SASP) that contributes to driving aging and loss of tissue homeostasis. The function of adult stem cell populations also reduces with age, contributing to the loss of tissue homeostasis. We previously demonstrated that injection of young, functional stem cells into mouse models of accelerated aging resulted in an extension of healthspan and lifespan through a cell non-autonomous mechanism(s). Interestingly, at least part of the ability of young, but not old stem cells to extend healthspan and lifespan in mice co-purifies with extracellular vesicles (EVs). EVs are comprised of microvesicles, released from the plasma membrane by shedding, and nanovesicles or exosomes. EVs are enriched for certain proteins, lipids, messenger RNA (mRNAs), and non-coding RNA (ncRNAs) including miRNAs, which can be transferred to target cells to modulate their function. We recently demonstrated that mesenchymal stem cell (MSC)-derived EVs can rescue markers of genotoxic stress-induced senescence in different cell types in culture and that senescent cells internalize EVs much more efficiently than control cells. We also demonstrated that injection of young MSC-derived EVs extends healthspan and lifespan in a mouse model of accelerated aging. Finally, we demonstrated that EVs isolated from the serum of young mice, but not old mice can suppress markers of senescence in both mouse and human cells as well as reduce senescence in vivo. Based on these results, we hypothesize that a subset(s) of circulating, blood-borne EVs can function as cell non-autonomous, anti-geronic factors through mechanisms involving modulation of cellular senescence and stem cell function as well as likely through other mechanisms. Loss of the production of these anti-geronic EVs contribute to aging. In addition, we hypothesize that subsets of circulating EVs, possibly derived from damaged, senescent cells, in old mice function as pro-geronic factors. Thus the goal of this proposal is to examine the novel geronic roles of the subset(s) of circulating geronic EVs from young and old mice in modulating aging, using young and old BM-MSC-derived and senescent and non-senescent MEF-derived EVs as controls. The successful completion of the proposed experiments will provide new insights into the mechanisms of aging and lead to novel approaches to extend healthspan.